Two tonoplast MATE proteins function as turgor-regulating chloride channels inArabidopsis

The central vacuole in a plant cell occupies the majority of the cellular volume and plays a key role in turgor regulation. The vacuolar membrane (tonoplast) contains a large number of transporters that mediate fluxes of solutes and water, thereby adjusting cell turgor in response to developmental and environmental signals. We report that two tonoplast Detoxification efflux carrier (DTX)/Multidrug and Toxic Compound Extrusion (MATE) transporters, DTX33 and DTX35, function as chloride channels essential for turgor regulation inArabidopsis. Ectopic expression of each transporter inNicotiana benthamianamesophyll cells elicited a large voltage-dependent inward chloride current across the tonoplast, showing that DTX33 and DTX35 each constitute a functional channel. Both channels are highly expressed inArabidopsistissues, including root hairs and guard cells that experience rapid turgor changes during root-hair elongation and stomatal movements. Disruption of these two genes, either in single or double mutants, resulted in shorter root hairs and smaller stomatal aperture, with double mutants showing more severe defects, suggesting that these two channels function additively to facilitate anion influx into the vacuole during cell expansion. In addition,dtx35single mutant showed lower fertility as a result of a defect in pollen-tube growth. Indeed, patch-clamp recording of isolated vacuoles indicated that the inward chloride channel activity across the tonoplast was impaired in the double mutant. Because MATE proteins are widely known transporters of organic compounds, finding MATE members as chloride channels expands the functional definition of this large family of transporters.

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Carotenoids in nature: insights from plants and beyond

Functional Plant Biology ◽

10.1071/fp11192 ◽

2011 ◽

Vol 38 (11) ◽

pp. 833 ◽

Cited By ~ 243

Author(s):

Christopher I. Cazzonelli

Keyword(s):

Biosynthetic Pathway ◽

Large Family ◽

New Wave ◽

Environmental Signals ◽

Carotenoid Accumulation ◽

Photosynthetic Organisms ◽

Essential Components ◽

Yellow Orange ◽

Commercially Important ◽

Bacteria And Fungi

Carotenoids are natural isoprenoid pigments that provide leaves, fruits, vegetables and flowers with distinctive yellow, orange and some reddish colours as well as several aromas in plants. Their bright colours serve as attractants for pollination and seed dispersal. Carotenoids comprise a large family of C40 polyenes and are synthesised by all photosynthetic organisms, aphids, some bacteria and fungi alike. In animals carotenoid derivatives promote health, improve sexual behaviour and are essential for reproduction. As such, carotenoids are commercially important in agriculture, food, health and the cosmetic industries. In plants, carotenoids are essential components required for photosynthesis, photoprotection and the production of carotenoid-derived phytohormones, including ABA and strigolactone. The carotenoid biosynthetic pathway has been extensively studied in a range of organisms providing an almost complete pathway for carotenogenesis. A new wave in carotenoid biology has revealed implications for epigenetic and metabolic feedback control of carotenogenesis. Developmental and environmental signals can regulate carotenoid gene expression thereby affecting carotenoid accumulation. This review highlights mechanisms controlling (1) the first committed step in phytoene biosynthesis, (2) flux through the branch to synthesis of α- and β-carotenes and (3) metabolic feedback signalling within and between the carotenoid, MEP and ABA pathways.

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A Highly Sensitive Cell-Based TLR Reporter Platform for the Specific Detection of Bacterial TLR Ligands

Frontiers in Immunology ◽

10.3389/fimmu.2021.817604 ◽

2022 ◽

Vol 12 ◽

Author(s):

Katharina Radakovics ◽

Claire Battin ◽

Judith Leitner ◽

Sabine Geiselhart ◽

Wolfgang Paster ◽

...

Keyword(s):

Limit Of Detection ◽

Ectopic Expression ◽

High Sensitivity ◽

High Specificity ◽

Reporter System ◽

Specificity And Sensitivity ◽

Highly Sensitive ◽

Definition Of ◽

The Individual ◽

Allergen Extracts

Toll-like receptors (TLRs) are primary pattern recognition receptors (PRRs), which recognize conserved microbial components. They play important roles in innate immunity but also in the initiation of adaptive immune responses. Impurities containing TLR ligands are a frequent problem in research but also for the production of therapeutics since TLR ligands can exert strong immunomodulatory properties even in minute amounts. Consequently, there is a need for sensitive tools to detect TLR ligands with high sensitivity and specificity. Here we describe the development of a platform based on a highly sensitive NF-κB::eGFP reporter Jurkat JE6-1 T cell line for the detection of TLR ligands. Ectopic expression of TLRs and their coreceptors and CRISPR/Cas9-mediated deletion of endogenously expressed TLRs was deployed to generate reporter cell lines selectively expressing functional human TLR2/1, TLR2/6, TLR4 or TLR5 complexes. Using well-defined agonists for the respective TLR complexes we could demonstrate high specificity and sensitivity of the individual reporter lines. The limit of detection for LPS was below 1 pg/mL and ligands for TLR2/1 (Pam3CSK4), TLR2/6 (Fsl-1) and TLR5 (flagellin) were detected at concentrations as low as 1.0 ng/mL, 0.2 ng/mL and 10 pg/mL, respectively. We showed that the JE6-1 TLR reporter cells have the utility to characterize different commercially available TLR ligands as well as more complex samples like bacterially expressed proteins or allergen extracts. Impurities in preparations of microbial compounds as well as the lack of specificity of detection systems can lead to erroneous results and currently there is no consensus regarding the involvement of TLRs in the recognition of several molecules with proposed immunostimulatory functions. This reporter system represents a highly suitable tool for the definition of structural requirements for agonists of distinct TLR complexes.

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A novel Xenopus mix-like gene milk involved in the control of the endomesodermal fates

Development ◽

10.1242/dev.125.14.2577 ◽

1998 ◽

Vol 125 (14) ◽

pp. 2577-2585 ◽

Cited By ~ 19

Author(s):

V. Ecochard ◽

C. Cayrol ◽

S. Rey ◽

F. Foulquier ◽

D. Caillol ◽

...

Keyword(s):

Marginal Zone ◽

Ectopic Expression ◽

Homeobox Gene ◽

Early Response ◽

Response Gene ◽

Mesodermal Cell ◽

Early Response Gene ◽

Mesoderm Formation ◽

Definition Of ◽

Early Gastrula

Here we describe a novel Xenopus homeobox gene, milk, related by sequence homology and expression pattern to the vegetally expressed Mix.1. As is the case with Mix.1, milk is an immediate early response gene to the mesoderm inducer activin. milk is expressed at the early gastrula stage in the vegetal cells, fated to form endoderm, and in the marginal zone fated to form mesoderm. During gastrulation, expression of milk becomes progressively reduced in the involuting mesodermal cells but is retained in the endoderm, suggesting that it may play a key role in the definition of the endo-mesodermal boundary in the embryo. Overexpression of milk in the marginal zone blocks mesodermal cell involution, represses the expression of several mesodermal genes such as Xbra, goosecoid, Xvent-1 or Xpo and increases the expression of the endodermal gene, endodermin. In the dorsal marginal zone, overexpression of milk leads to a severe late phenotype including the absence of axial structures. Ectopic expression of milk in the animal hemisphere or in ectodermal explants induces a strong expression of endodermin. Taken together, we propose that milk plays a role in the correct patterning of the embryo by repressing mesoderm formation and promoting endoderm identity.

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The foundations of plant intelligence

Interface Focus ◽

10.1098/rsfs.2016.0098 ◽

2017 ◽

Vol 7 (3) ◽

pp. 20160098 ◽

Cited By ~ 40

Author(s):

Anthony Trewavas

Keyword(s):

Single Cells ◽

Wild Plants ◽

Neural Systems ◽

Plant Interactions ◽

Distributed Resources ◽

Environmental Signals ◽

Plant Intelligence ◽

Sense And Respond ◽

Definition Of ◽

Plant Behaviour

Intelligence is defined for wild plants and its role in fitness identified. Intelligent behaviour exhibited by single cells and systems similarity between the interactome and connectome indicates neural systems are not necessary for intelligent capabilities. Plants sense and respond to many environmental signals that are assessed to competitively optimize acquisition of patchily distributed resources. Situations of choice engender motivational states in goal-directed plant behaviour; consequent intelligent decisions enable efficient gain of energy over expenditure. Comparison of swarm intelligence and plant behaviour indicates the origins of plant intelligence lie in complex communication and is exemplified by cambial control of branch function. Error correction in behaviours indicates both awareness and intention as does the ability to count to five. Volatile organic compounds are used as signals in numerous plant interactions. Being complex in composition and often species and individual specific, they may represent the plant language and account for self and alien recognition between individual plants. Game theory has been used to understand competitive and cooperative interactions between plants and microbes. Some unexpected cooperative behaviour between individuals and potential aliens has emerged. Behaviour profiting from experience, another simple definition of intelligence, requires both learning and memory and is indicated in the priming of herbivory, disease and abiotic stresses.

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Ethylene promotes root hair growth through coordinated EIN3/EIL1 and RHD6/RSL1 activity in Arabidopsis

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1711723115 ◽

2017 ◽

Vol 114 (52) ◽

pp. 13834-13839 ◽

Cited By ~ 42

Author(s):

Ying Feng ◽

Ping Xu ◽

Bosheng Li ◽

Pengpeng Li ◽

Xing Wen ◽

...

Keyword(s):

Root Hair ◽

Hair Growth ◽

Root Hairs ◽

Environmental Signals ◽

Environmental Interactions ◽

Two Factors ◽

Root Hair Growth ◽

Transcriptional Complex ◽

Root Hair Initiation ◽

Molecular Framework

Root hairs are an extensive structure of root epidermal cells and are critical for nutrient acquisition, soil anchorage, and environmental interactions in sessile plants. The phytohormone ethylene (ET) promotes root hair growth and also mediates the effects of different signals that stimulate hair cell development. However, the molecular basis of ET-induced root hair growth remains poorly understood. Here, we show that ET-activated transcription factor ETHYLENE-INSENSITIVE 3 (EIN3) physically interacts with ROOT HAIR DEFECTIVE 6 (RHD6), a well-documented positive regulator of hair cells, and that the two factors directly coactivate the hair length-determining gene RHD6-LIKE 4 (RSL4) to promote root hair elongation. Transcriptome analysis further revealed the parallel roles of the regulator pairs EIN3/EIL1 (EIN3-LIKE 1) and RHD6/RSL1 (RHD6-LIKE 1). EIN3/EIL1 and RHD6/RSL1 coordinately enhance root hair initiation by selectively regulating a subset of core root hair genes. Thus, our work reveals a key transcriptional complex consisting of EIN3/EIL1 and RHD6/RSL1 in the control of root hair initiation and elongation, and provides a molecular framework for the integration of environmental signals and intrinsic regulators in modulating plant organ development.

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ClC-K chloride channels: emerging pathophysiology of Bartter syndrome type 3

AJP Renal Physiology ◽

10.1152/ajprenal.00004.2015 ◽

2015 ◽

Vol 308 (12) ◽

pp. F1324-F1334 ◽

Cited By ~ 32

Author(s):

Olga Andrini ◽

Mathilde Keck ◽

Rodolfo Briones ◽

Stéphane Lourdel ◽

Rosa Vargas-Poussou ◽

...

Keyword(s):

Chloride Channel ◽

Chloride Channels ◽

Large Family ◽

Functional Expression ◽

Bartter Syndrome ◽

Channel Structure ◽

Syndrome Type ◽

Missense Mutations ◽

Membrane Expression ◽

Type 3

The mutations in the CLCNKB gene encoding the ClC-Kb chloride channel are responsible for Bartter syndrome type 3, one of the four variants of Bartter syndrome in the genetically based nomenclature. All forms of Bartter syndrome are characterized by hypokalemia, metabolic alkalosis, and secondary hyperaldosteronism, but Bartter syndrome type 3 has the most heterogeneous presentation, extending from severe to very mild. A relatively large number of CLCNKB mutations have been reported, including gene deletions and nonsense or missense mutations. However, only 20 CLCNKB mutations have been functionally analyzed, due to technical difficulties regarding ClC-Kb functional expression in heterologous systems. This review provides an overview of recent progress in the functional consequences of CLCNKB mutations on ClC-Kb chloride channel activity. It has been observed that 1) all ClC-Kb mutants have an impaired expression at the membrane; and 2) a minority of the mutants combines reduced membrane expression with altered pH-dependent channel gating. Although further investigation is needed to fully characterize disease pathogenesis, Bartter syndrome type 3 probably belongs to the large family of conformational diseases, in which the mutations destabilize channel structure, inducing ClC-Kb retention in the endoplasmic reticulum and accelerated channel degradation.

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Ectopic Expression of an Activated RAC inArabidopsisDisrupts Membrane Cycling

Molecular Biology of the Cell ◽

10.1091/mbc.e04-07-0562 ◽

2005 ◽

Vol 16 (4) ◽

pp. 1913-1927 ◽

Cited By ~ 88

Author(s):

Daria Bloch ◽

Meirav Lavy ◽

Yael Efrat ◽

Idan Efroni ◽

Keren Bracha-Drori ◽

...

Keyword(s):

Transgenic Plants ◽

Actin Cytoskeleton ◽

Protein Trafficking ◽

Rho Gtpases ◽

Fluorescent Protein ◽

Ectopic Expression ◽

Root Hairs ◽

Brefeldin A ◽

Cytoskeleton Organization ◽

Trafficking Inhibitor

Rho GTPases regulate the actin cytoskeleton, exocytosis, endocytosis, and other signaling cascades. Rhos are subdivided into four subfamilies designated Rho, Racs, Cdc42, and a plant-specific group designated RACs/Rops. This research demonstrates that ectopic expression of a constitutive active Arabidopsis RAC, AtRAC10, disrupts actin cytoskeleton organization and membrane cycling. We created transgenic plants expressing either wild-type or constitutive active AtRAC10 fused to the green fluorescent protein. The activated AtRAC10 induced deformation of root hairs and leaf epidermal cells and was primarily localized in Triton X-100–insoluble fractions of the plasma membrane. Actin cytoskeleton reorganization was revealed by creating double transgenic plants expressing activated AtRAC10 and the actin marker YFP-Talin. Plants were further analyzed by membrane staining with N-[3-triethylammoniumpropyl]-4-[p-diethylaminophenylhexatrienyl] pyridinium dibromide (FM4-64) under different treatments, including the protein trafficking inhibitor brefeldin A or the actin-depolymeryzing agents latrunculin-B (Lat-B) and cytochalasin-D (CD). After drug treatments, activated AtRAC10 did not accumulate in brefeldin A compartments, but rather reduced their number and colocalized with FM4-64–labeled membranes in large intracellular vesicles. Furthermore, endocytosis was compromised in root hairs of activated AtRAC10 transgenic plants. FM4-64 was endocytosed in nontransgenic root hairs treated with the actin-stabilizing drug jasplakinolide. These findings suggest complex regulation of membrane cycling by plant RACs.

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Binary sequences and lattices constructed by discrete logarithms

AIMS Mathematics ◽

10.3934/math.2022259 ◽

2021 ◽

Vol 7 (3) ◽

pp. 4655-4671

Author(s):

Yuchan Qi ◽

◽

Huaning Liu

Keyword(s):

Exponential Sums ◽

Binary Sequence ◽

Large Family ◽

Binary Sequences ◽

Multiplicative Inverse ◽

Discrete Logarithms ◽

Modulo P ◽

Large Families ◽

Definition Of ◽

Pseudorandom Measures

<abstract><p>In 1997, Mauduit and Sárközy first introduced the measures of pseudorandomness for binary sequences. Since then, many pseudorandom binary sequences have been constructed and studied. In particular, Gyarmati presented a large family of pseudorandom binary sequences using the discrete logarithms. Ten years later, to satisfy the requirement from many applications in cryptography (e.g., in encrypting "bit-maps'' and watermarking), the definition of binary sequences is extended from one dimension to several dimensions by Hubert, Mauduit and Sárközy. They introduced the measure of pseudorandomness for this kind of several-dimension binary sequence which is called binary lattices. In this paper, large families of pseudorandom binary sequences and binary lattices are constructed by both discrete logarithms and multiplicative inverse modulo $ p $. The upper estimates of their pseudorandom measures are based on estimates of either character sums or mixed exponential sums.</p></abstract>

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The Ca2+ sensor protein CMI1 fine tunes root development, auxin distribution and responses

10.1101/488536 ◽

2018 ◽

Author(s):

Ora Hazak ◽

Elad Mamon ◽

Meirav Lavy ◽

Hasana Sternberg ◽

Smrutisanjita Behera ◽

...

Keyword(s):

Root Growth ◽

Root Development ◽

Lateral Root ◽

Developmental Plasticity ◽

Molecular Mechanisms ◽

Ectopic Expression ◽

Growth Arrest ◽

Root Hairs ◽

Root Tip ◽

Auxin Distribution

Signaling cross-talks between auxin, a regulator of plant development and Ca2+, a universal second messenger have been proposed to modulate developmental plasticity in plants. However, the underlying molecular mechanisms are largely unknown. Here we report that in Arabidopsis roots, auxin elicits specific Ca2+ signaling pattern that spatially coincide with the expression pattern of auxin-regulated genes. We identified the EF-hand protein CMI1 (Ca2+ sensor Modulator of ICR1) as an interactor of the ROP effector ICR1 (Interactor of Constitutively active ROP). CMI1 is monomeric in solution, changes its secondary structure at Ca2+ concentrations ranging from 10-9 to 10-8 M and its interaction with ICR1 is Ca2+ dependent, involving a conserved hydrophobic pocket. cmi1 mutants display an increased auxin response including shorter primary roots, longer root hairs, longer hypocotyls and altered lateral root formation while ectopic expression of CMI1 induces root growth arrest and reduced auxin responses at the root tip. When expressed alone, CMI1 is localized at the plasma membrane, the cytoplasm and in nuclei. Interaction of CMI1 and ICR1 results in exclusion of CMI1 from nuclei and suppression of the root growth arrest. CMI1 expression is directly upregulated by auxin while expression of auxin induced genes is enhanced in cmi1 concomitantly with repression of auxin induced Ca2+ increases in the lateral root cap and vasculature, indicating that CMI1 represses early auxin responses. Collectively, our findings identify a crucial function of Ca2+ signaling and CMI1 in root growth and suggest an auxin-Ca2+ regulatory feedback loop that fine tunes root development.

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Developmental regulation of a cyclin-dependent kinase inhibitor controls postembryonic cell cycle progression in Caenorhabditis elegans

Development ◽

10.1242/dev.125.18.3585 ◽

1998 ◽

Vol 125 (18) ◽

pp. 3585-3597 ◽

Cited By ~ 23

Author(s):

Y. Hong ◽

R. Roy ◽

V. Ambros

Keyword(s):

Cell Cycle ◽

Cell Cycle Progression ◽

Kinase Inhibitors ◽

Kinase Inhibitor ◽

Developmental Regulation ◽

Ectopic Expression ◽

Cyclin Dependent Kinase ◽

Cycle Progression ◽

Environmental Signals ◽

C Elegans

C. elegans cki-1 encodes a member of the CIP/KIP family of cyclin-dependent kinase inhibitors, and functions to link postembryonic developmental programs to cell cycle progression. The expression pattern of cki-1::GFP suggests that cki-1 is developmentally regulated in blast cells coincident with G1, and in differentiating cells. Ectopic expression of CKI-1 can prematurely arrest cells in G1, while reducing cki-1 activity by RNA-mediated interference (RNAi) causes extra larval cell divisions, suggesting a role for cki-1 in the developmental control of G1/S. cki-1 activity is required for the suspension of cell cycling that occurs in dauer larvae and starved L1 larvae in response to environmental signals. In vulva precursor cells (VPCs), a pathway of heterochronic genes acts via cki-1 to maintain VPCs in G1 during the L2 stage.

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